Project Summary Lung cancer is the most common cause of cancer mortality in the US and worldwide. With limited effective treatments, early detection and surgical resection remain the treatment of choice. Unfortunately, at the time of diagnosis only 15% of patients with lung cancer have localized resectable disease. Advances in low-dose computed tomography (CT) screening now allow lung nodules to be detected early, and have increased survival by 20%, but low dose CT cannot distinguish the small number of malignant nodules from the majority of benign ones. Therefore a simple, non-invasive test that can accurately distinguish the malignant from benign nodules remains an important clinical goal. To identify candidate biomarkers for NSCLC, , we analyzed RNA from the peripheral blood mononuclear cell (PBMC) fraction of whole blood from patients with non-small cell lung cancer (NSCLC) and controls, using unbiased forward genetic screening approaches. We identified AKAP4 as a highly accurate blood biomarker for the presence of NSCLC. The area under receiver operating characteristics curve (AUC) is 0.9714 (sensitivity and specificity are 92.80% and 92.59%, respectively) when blood samples from 264 NSCLC patients were compared with 135 controls A separate analysis of only the 136 Stage I NSCLC cases improved the AUC to 0.9790, and a comparison of NSCLC patient samples to samples from 27 patients with high risk but histologically confirmed benign lung nodules gave an AUC of 0.9845, showing that AKAP4 expression in blood may be an excellent biomarker for early stage lung cancer and differentiation of malignant from benign nodules. To validate the accuracy of AKAP4 as a blood biomarker for NSCLC in a larger cohort, laying the groundwork for further development for clinical use, we propose the following specific aims: Specific Aim 1: Validate AKAP4 as a blood biomarker for the detection of early stage non-small cell lung cancer; test AKAP4 as a marker of recurrence. A. AKAP4 expression will be determined in PBMC samples from 500 Stage 1 and 2 NSCLC and 500 disease-free smokers and ex-smokers to assess its accuracy in detecting early stage NSCLC. B. Expanding on preliminary studies, AKAP4 expression will be assayed in PBMC samples from 374 NSCLC patients with longitudinal blood and CT data to determine its utility and accuracy in assessing remission and predicting recurrence. Specific Aim 2: Validate AKAP4 as a marker for malignant pulmonary nodules and determine the cellular source of AKAP4 expression. A. AKAP4 expression will be determined in PBMC samples from 500 NSCLC patients and 500 patients with benign lung nodules to assess whether AKAP4 expression can distinguish malignant from benign pulmonary nodules. B. Determine the cellular source of AKAP4 expression. We will determine the AKAP4 expression in 12 types of human blood cells, as AKAP4 may derive from circulating tumor cells or from immune cells.